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[This corrects the article DOI: 10.1371/journal.pone.0248810.].
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Background: This study aimed to investigate the effects of a collagen endometrial patch (EM patch) loaded with adipose-derived mesenchymal stem cells (ADSCs) on endometrial regeneration in a rat model with thin endometrium. Materials and methods: Thin endometrium was induced in female rats and divided into treatment groups as outlined: control, group 1(G1), local injection of ADSCs into the uterus, group 2 (G2), an EM patch without ADSCs, group 3 (G3), and an EM patch loaded with ADSCs, group 4 (G4). The rats were euthanized at either two weeks or four weeks after modeling and treatment followed by histological and biochemical analyses to examine the regenerative effects on the injured endometrium. Results: Transplantation of the ADSC-loaded EM patch significantly promoted endometrial proliferation and increased the luminal epithelial area. Two weeks after treatment, the mean number of von Villebrand factor (vWF)+ or cluster of differentiation (CD) 31+-stained blood vessels was significantly higher in G4 than in G1 and G2. The mRNA and protein expression levels of TGF-ß and FGF2 were significantly upregulated in G4 compared to those in the control. G4 exhibited significantly increased LIF mRNA levels and immunoreactivity compared with the other groups at both two weeks and four weeks after treatment. Cell tracking after ADSCs treatment revealed the presence of a substantial number of ADSCs grafted in the uterine tissues of G4, whereas a low number of ADSCs that were focally clustered were present in G2. Conclusion: Transplantation of EM patches loaded with ADSCs resulted in the histological and biochemical restoration of an injured endometrium. The strategic integration of EM patches and ADSCs holds significant promise as an innovative therapeutic approach for effectively treating impaired endometrial conditions.
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Células Madre Mesenquimatosas , Regeneración , Ratas , Femenino , Animales , Ratas Sprague-Dawley , Endometrio/patología , Colágeno/metabolismo , ARN Mensajero/metabolismoRESUMEN
A disintegrin and metalloprotease 10 (ADAM10) regulates the expression of cell surface receptors such as tumor necrosis factor receptor 1, toll-like receptor 4, and the receptor for advanced glycation end products (RAGE) by cleaving their extracellular regions. To function as a sheddase, ADAM10 should translocate from the intracellular compartments to the cell surface, but the translocation mechanism remains unclear. In this study, we explored the possible role of adenosine monophosphate-activated protein kinase (AMPK) in the induction of ADAM10 shedding activity. In cultured human aortic endothelial cells (HAECs), 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR), an AMPK activator, boosted ADAM10 cell surface translocation and ectodomain shedding of RAGE. ADAM10 inhibition with GI 254023X and ADAM10 siRNA silencing both prevented AICAR-induced RAGE ectodomain shedding. AICAR increased AMPK phosphorylation as well. Both Compound C-mediated AMPK inhibition and AMPKα1-siRNA-mediated AMPK depletion suppressed AICAR-induced ADAM10 cell surface translocation and RAGE ectodomain shedding. On the other hand, siRNA knockdown of Rab14, a small GTPase that facilitates the intracellular trafficking of transmembrane proteins, prevented AICAR-induced ADAM10 cell surface translocation and RAGE ectodomain shedding. In conclusion, AMPK activation is an obvious inducer of ADAM10 shedding activity. Our findings suggest that AMPK boosts ADAM10 shedding activity in HAECs by promoting Rab14-dependent ADAM10 cell surface translocation.
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Proteínas Quinasas Activadas por AMP , Células Endoteliales , Humanos , Células Endoteliales/metabolismo , Proteínas Quinasas Activadas por AMP/metabolismo , Proteína ADAM10/metabolismo , Membrana Celular/metabolismo , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismo , Receptor para Productos Finales de Glicación Avanzada/metabolismo , Secretasas de la Proteína Precursora del Amiloide/metabolismo , Proteínas de la Membrana/metabolismo , Proteínas de Unión al GTP rab/metabolismoRESUMEN
To overcome the limitations of lymphedema treatment, human adipose-derived stem cells (hADSCs) were injected into decellularized lymph nodes to produce a recellularized lymph node-scaffold, and the effect of lymphangiogenesis was investigated in lymphedema animal models. Axillary lymph nodes were harvested from Sprague Dawley rats (7 weeks old, 220-250 g) for decellularization. The decellularized lymph nodes were performed and PKH26-labeled hADSCs (1 × 106/50 µL) were injected in the decellularized lymph node-scaffolds. Forty rats were equally divided into four groups: lymphedema as control group, hADSC group, decellularized lymph node-scaffold group, and recellularized lymph node-scaffold group. The lymphedema model was made by removing inguinal lymph nodes, and hADSCs or scaffolds were transplanted. Histopathological assessments were performed by hematoxylin and eosin and Masson's trichrome staining. Lymphangiogenesis was evaluated by Immunofluorescence staining and western blot. Decellularized lymph nodes showed virtually complete absence of cellular material and maintenance of lymph node architecture. The hADSCs were significantly observed in recellularized lymph node-scaffolds group. The recellularized lymph node-scaffold group was histologically similar to normal lymph nodes. The vascular endothelial growth factor A and lymphatic vessel endothelial hyaluronan receptor 1 (LYVE-1) in immunofluorescence staining were highly expressed in recellularized lymph node-scaffolds group. Also, the expression of LYVE-1 protein significantly increased in recellularized lymph node-scaffold group compared with others. Recellularized lymph node -scaffold had a much better therapeutic effect than stem cells or decellularized lymph node-scaffold alone, and induced stable lymphangiogenesis.
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Linfedema , Factor A de Crecimiento Endotelial Vascular , Humanos , Ratas , Animales , Factor A de Crecimiento Endotelial Vascular/farmacología , Ratas Sprague-Dawley , Ganglios Linfáticos , Linfedema/terapia , Regeneración , Células Madre , Andamios del TejidoRESUMEN
AIMS: Micromotion is an autonomous intramural movement of the bladder, and is believed to be an initial step in the generation of urinary urgency. Therefore, controlling micromotion may be a novel target in overactive bladder (OAB) treatment. However, developing micromotion treatment has been limited by the absence of a standardized animal model. We attempted to create a micromotion animal model and investigated the effectiveness of a ß3 -adrenoceptor agonist (CL316,243) on micromotion. METHODS: Bilateral major pelvic ganglia (MPGs) were excised in 18 male Sprague-Dawley rats, resulting in an almost completely denervated bladder. On postoperative Day 7, cystometry was performed. Rats were divided into three treatment groups: CL316,243; ß3- adrenoceptor antagonist (SR59230A) pretreated CL316,243; and a nonselective antimuscarinic agent (oxybutynin). Changes in micromotion were evaluated after the intra-arterial administration of each agent. RESULTS: Low-amplitude oscillations in intravesical pressure (micromotion) were observed 1 week after MPGs excision. Micromotion frequency significantly (p = 0.003) decreased (2.17 ± 3.54 times/5 min) with CL316,243 compared with vehicle (6.33 ± 1.97 times/5 min). Micromotion amplitude also decreased with CL316,243 (1.15 ± 1.93 cmH2 O) compared with vehicle (5.96 ± 5.12 cmH2 O), approaching conventional significance (p = 0.090). No significant decreases in frequency or amplitude were observed with oxybutynin treatment. CONCLUSIONS: Systemic administration of the ß3 -adrenoceptor agonist CL316,243 effectively controlled micromotion in bilateral MPGs-excised, almost completely denervated rat bladders. This result indicates that ß3 -adrenoceptor agonist may affect the bladder directly, suggesting that it might be effective for overall OAB, regardless of the presence or level of neurological deficits. Bilateral MPGs-excised rats are considered a plausible micromotion animal model suitable for future research.
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Vejiga Urinaria Hiperactiva , Vejiga Urinaria , Animales , Masculino , Ratas , Agonistas de Receptores Adrenérgicos beta 3/farmacología , Ratas Sprague-Dawley , Receptores Adrenérgicos , Receptores Adrenérgicos beta 3RESUMEN
BACKGROUND: Metabolic rewiring in microbes is an economical and sustainable strategy for synthesizing valuable natural terpenes. Terpenes are the largest class of nature-derived specialized metabolites, and many have valuable pharmaceutical or biological activity. Squalene, a medicinal terpene, is used as a vaccine adjuvant to improve the efficacy of vaccines, including pandemic coronavirus disease 2019 (COVID-19) vaccines, and plays diverse biological roles as an antioxidant and anticancer agent. However, metabolic rewiring interferes with inherent metabolic pathways, often in a way that impairs the cellular growth and fitness of the microbial host. In particular, as the key starting molecule for producing various compounds including squalene, acetyl-CoA is involved in numerous biological processes with tight regulation to maintain metabolic homeostasis, which limits redirection of metabolic fluxes toward desired products. RESULTS: In this study, focusing on the recycling of surplus metabolic energy stored in lipid droplets, we show that the metabolic recycling of the surplus energy to acetyl-CoA can increase squalene production in yeast, concomitant with minimizing the metabolic interferences in inherent pathways. Moreover, by integrating multiple copies of the rate-limiting enzyme and implementing N-degron-dependent protein degradation to downregulate the competing pathway, we systematically rewired the metabolic flux toward squalene, enabling remarkable squalene production (1024.88 mg/L in a shake flask). Ultimately, further optimization of the fed-batch fermentation process enabled remarkable squalene production of 6.53 g/L. CONCLUSIONS: Our demonstration of squalene production via engineered yeast suggests that plant- or animal-based supplies of medicinal squalene can potentially be complemented or replaced by industrial fermentation. This approach will also provide a universal strategy for the more stable and sustainable production of high-value terpenes.
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Myelodysplastic syndrome (MDS) is a heterogeneous hematopoietic disorder associated with cellular proliferative and apoptotic activity. We retrospectively investigated these activities in bone marrow samples from 76 MDS patients using immunohistochemical staining for Ki-67 and cleaved caspase-3. We divided cleaved caspase-3 into two groups based on median value and compared the differences according to MDS risk scoring systems. We compared MDS patient indices with idiopathic cytopenia of undetermined significance (ICUS) and healthy control (HC) indices using our previously published data. Cleaved caspase-3 immunohistochemistry was highest in MDS patients, followed by ICUS patients and HCs. Similarly, the mean Ki-67 grade was also highest in MDS patients, followed by ICUS patients and HCs. Higher cleaved caspase-3 grade was significantly associated with lower IPSS-R score (p = 0.020), whereas Ki-67 was not associated with MDS. Interestingly, TET2 mutation was associated with decreased cleaved caspase-3 levels (p = 0.03). However, there was no significant association between proliferative/apoptotic activity and survival. Our results suggest that apoptotic activity gradually increases from healthy controls and ICUS patients to MDS patients. Furthermore, higher apoptotic activity was associated with better MDS patient prognostic scores. Further studies are needed to reveal the differences in apoptotic activity between lower- and higher-risk MDS.
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AIMS: Glucagon-like peptide-1 alleviates the deleterious effects of advanced glycation end products (AGEs), but the underlying mechanisms are not fully understood. In this study, we investigated the protective mechanism using liraglutide, a glucagon-like peptide-1 receptor agonist, in cultured human aortic endothelial cells (HAECs). MAIN METHODS: Following liraglutide treatment in HAECs, the receptor for AGEs (RAGE) was measured in both cell lysate and culture supernatant, the cytosolic free Ca2+ level was monitored using Fluo-4 AM, the phosphorylation of adenosine monophosphate-activated protein kinase (AMPK) was analyzed, and immunofluorescence staining was used to visualize a disintegrin and metalloprotease 10 (ADAM10) on the cell surface. KEY FINDINGS: Liraglutide (100 nM) induced ectodomain shedding of RAGE within 30 min and inhibited the expression of intercellular adhesion molecule-1 (ICAM-1) induced by AGEs of bovine serum albumin (AGE-BSA). Further experiments revealed that liraglutide rapidly increases extracellular Ca2+ influx through L-type calcium channels and activates AMPK, resulting in the translocation of ADAM10 to the cell surface, whereas siRNA-mediated ADAM10 depletion prevents liraglutide-induced ectodomain shedding of RAGE and eliminates liraglutide's inhibitory effect on AGE-BSA-induced ICAM-1 expression. Moreover, compound C-mediated AMPK inhibition and siRNA-mediated AMPK depletion both prevented ADAM10 translocation to the cell surface and ADAM10-mediated ectodomain shedding of RAGE. SIGNIFICANCE: Liraglutide reduces the number of intact RAGE on the cell surface by inducing ADAM10-mediated ectodomain shedding, which decreases the inflammatory effects of AGEs. AMPK activated by extracellular Ca2+ influx is critically involved in the translocation of ADAM10 to the cell surface, where it cleaves RAGE.
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Aorta , Células Epiteliales , Receptor del Péptido 1 Similar al Glucagón/antagonistas & inhibidores , Productos Finales de Glicación Avanzada/metabolismo , Liraglutida/farmacología , Aorta/efectos de los fármacos , Aorta/patología , Línea Celular , Células Epiteliales/efectos de los fármacos , Células Epiteliales/patología , HumanosRESUMEN
BACKGROUND/AIMS: Chronic intestinal pseudo-obstruction (CIPO) is a clinically heterogeneous syndrome characterized by compromised peristalsis and intestinal obstruction. Variants of actin gamma 2 (ACTG2), a protein crucial for correct enteric muscle contraction, have been found in CIPO patients. The aim of this study is to examine the clinical features and ACTG2 variants in Korean patients with CIPO. METHODS: From January 1995 to August 2020, 12 patients diagnosed with CIPO were included and genetic analysis testing of ACTG2 was performed. RESULTS: Heterozygous ACTG2 missense variants were found in 6 patients (50.0%). The p.Arg257Cys variant was found in 3 patients, and p.Arg63Gln and p.Arg178His variants were found in 1 patient each. A novel variant, p.Ile193Phe, was found in 1 patient. Three patients were diagnosed at birth, 2 at the age of 1 year, and 1 at 3 years of age. Abnormal prenatal genitourinary ultrasonographic findings were found in all 6 patients; microcolon was found in 4 patients (66.7%), and megacystis in all 6 patients. The pathology showed abnormal ganglion cells as well as myopathic findings. All patients are dependent on total parenteral nutrition and are to date alive. CONCLUSIONS: ACTG2 variants are commonly found in Korean patients with CIPO. In CIPO patients with megacystis and abnormal prenatal ultrasonography, genetic testing of ACTG2 should be considered. Molecular diagnosis of CIPO is more important than pathologic diagnosis.
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BACKGROUND: Inflammatory bowel disease (IBD) is associated with an increased risk of Colorectal cancer (CRC), and its most important risk factors are the duration and extent of the disease. Pediatric-onset inflammatory bowel disease has a tendency for a more extensive, more severe, and longer predicted disease duration than adult-onset inflammatory bowel disease. This study aimed to identify the clinical characteristics of patients with CRC related to pediatric-onset IBD and consider the appropriateness of current surveillance endoscopy recommendations for the detection of premalignant lesions and early-stage CRC. METHODS: We searched a research platform based on the SUPREME electronic medical record data-mining system to identify cases of colorectal malignancy in patients with pediatric IBD that presented between 2000 and 2020. RESULTS: During the follow-up, 4 (1.29 per 1000 person years) out of 443 patients with PIBD was diagnosed with CRC. The median age at diagnosis of CRC was 18.5 (range: 15-24) years, and the median period from diagnosis of IBD to CRC was 9.42 (range: 0.44-11.96) years. The sigmoid colon was the most frequent location of CRC (in 3 of the 4 cases). Adenocarcinoma was the most common histological type (in 2 of the 4 cases). CONCLUSIONS: Patients with pediatric-onset IBD exhibited a much shorter disease duration than that of adult-onset IBD at the time of diagnosis of CRC, suggesting that surveillance endoscopy for the detection of precancerous lesions and early-stage cancer should be initiated earlier in pediatric patients than in adult patients.
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Colitis Ulcerosa , Neoplasias Colorrectales , Enfermedad de Crohn , Enfermedades Inflamatorias del Intestino , Niño , Neoplasias Colorrectales/complicaciones , Neoplasias Colorrectales/diagnóstico , Neoplasias Colorrectales/epidemiología , Enfermedad de Crohn/complicaciones , Enfermedad de Crohn/diagnóstico , Humanos , Enfermedades Inflamatorias del Intestino/complicaciones , Enfermedades Inflamatorias del Intestino/diagnóstico , Factores de RiesgoRESUMEN
INTRODUCTION: Chronic inflammation and impaired neovascularization play critical roles in delayed wound healing in diabetic patients. To overcome the limitations of current diabetic wound (DBW) management interventions, we investigated the effects of a catechol-functionalized hyaluronic acid (HA-CA) patch combined with adipose-derived mesenchymal stem cells (ADSCs) in DBW mouse models. METHODS: Diabetes in mice (C57BL/6, male) was induced by streptozotocin (50 mg/kg, >250 mg/dL). Mice were divided into four groups: control (DBW) group, ADSCs group, HA-CA group, and HA-CA + ADSCs group (n = 10 per group). Fluorescently labeled ADSCs (5 × 105 cells/100 µL) were transplanted into healthy tissues at the wound boundary or deposited at the HA-CA patch at the wound site. The wound area was visually examined. Collagen content, granulation tissue thickness and vascularity, cell apoptosis, and re-epithelialization were assessed. Angiogenesis was evaluated by immunohistochemistry, quantitative real-time polymerase chain reaction, and Western blot. RESULTS: DBW size was significantly smaller in the HA-CA + ADSCs group (8% ± 2%) compared with the control (16% ± 5%, p < 0.01) and ADSCs (24% ± 17%, p < 0.05) groups. In mice treated with HA-CA + ADSCs, the epidermis was regenerated, and skin thickness was restored. CD31 and von Willebrand factor-positive vessels were detected in mice treated with HA-CA + ADSCs. The mRNA and protein levels of VEGF, IGF-1, FGF-2, ANG-1, PIK, and AKT in the HA-CA + ADSCs group were the highest among all groups, although the Spred1 and ERK expression levels remained unchanged. CONCLUSIONS: The combination of HA-CA and ADSCs provided synergistic wound healing effects by maximizing paracrine signaling and angiogenesis via the PI3K/AKT pathway. Therefore, ADSC-loaded HA-CA might represent a novel strategy for the treatment of DBW.
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Tejido Adiposo/metabolismo , Vendajes , Diabetes Mellitus Experimental/terapia , Angiopatías Diabéticas/terapia , Ácido Hialurónico , Trasplante de Células Madre , Células Madre/metabolismo , Cicatrización de Heridas , Heridas y Lesiones/terapia , Tejido Adiposo/patología , Animales , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patología , Angiopatías Diabéticas/metabolismo , Angiopatías Diabéticas/patología , Femenino , Humanos , Ácido Hialurónico/química , Ácido Hialurónico/farmacología , Masculino , Ratones , Células Madre/patología , Heridas y Lesiones/metabolismo , Heridas y Lesiones/patologíaRESUMEN
BACKGROUND: In Korea, there were issues regarding the use of immunoassays for anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibodies to detect infection. So, we compared antibody results of eight kinds of commercial immunoassays using clinical remnant specimens. METHODS: We compared the results of several immunoassay kits tested on 40 serum samples from 15 confirmed patients and 86 remnant serum samples from clinical laboratory. Eight kinds of IVD kits-four enzyme-linked immunosorbent assay, two lateral flow rapid immunochromatographic assays, and two chemiluminescent immunoassays with one RUO kit were tested. RESULTS: Among 40 serum samples from 15 coronavirus disease 2019 (COVID-19) patients, 35 yielded at least one positive result for detecting antibodies in the combined assessment. There were inconsistent results in 12 (28%) samples by single immunoassay. Forty samples collected in 2019 before the first COVID-19 Korean case showed negative results except for one equivocal result. CONCLUSION: The discrepant results obtained with different immunoassay kits in this study show that serological assessment of SARS-CoV-2 by a single immunoassay requires caution not only in detecting infection but also in assessing immunologic status.
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Anticuerpos Antivirales/sangre , COVID-19/diagnóstico , Inmunoensayo/métodos , SARS-CoV-2/inmunología , COVID-19/virología , Hospitalización , Humanos , Inmunoglobulina G/sangre , Juego de Reactivos para Diagnóstico , SARS-CoV-2/aislamiento & purificaciónRESUMEN
BACKGROUND: The effect of alcohol on the outcome and fibrinolysis phenotype in trauma patients remains unclear. Hence, we performed this study to determine whether alcohol is a risk factor for mortality and fibrinolysis shutdown in trauma patients. MATERIALS AND METHODS: A total of 686 patients who presented to our trauma center and underwent rotational thromboelastometry were included in the study. The primary outcome was in-hospital mortality. Logistic regression analysis was performed to determine whether alcohol was an independent risk factor for in-hospital mortality and fibrinolysis shutdown. RESULTS: The rate of in-hospital mortality was 13.8% and blood alcohol was detected in 27.7% of the patients among our study population. The patients in the alcohol-positive group had higher mortality rate, higher clotting time, and lower maximum lysis, more fibrinolysis shutdown, and hyperfibrinolysis than those in the alcohol-negative group. In logistic regression analysis, blood alcohol was independently associated with in-hospital mortality (odds ratio [OR] 2.578; 95% confidence interval [CI], 1.550-4.288) and fibrinolysis shutdown (OR 1.883 [95% CI, 1.286-2.758]). Within the fibrinolysis shutdown group, blood alcohol was an independent predictor of mortality (OR 2.168 [95% CI, 1.030-4.562]). CONCLUSIONS: Alcohol is an independent risk factor for mortality and fibrinolysis shutdown in trauma patients. Further, alcohol is an independent risk factor for mortality among patients who experienced fibrinolysis shutdown.
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Intoxicación Alcohólica/complicaciones , Coagulación Sanguínea , Fibrinólisis , Heridas y Lesiones/sangre , Heridas y Lesiones/mortalidad , Adulto , Anciano , Etanol/sangre , Femenino , Mortalidad Hospitalaria , Humanos , Modelos Logísticos , Masculino , Persona de Mediana Edad , FenotipoRESUMEN
INTRODUCTION: Remote ischemic preconditioning (rIPC) is a preventive strategy against ischemia-reperfusion injury. To reduce ischemia-reperfusion injury of random-pattern skin flaps, we investigated the therapeutic effects of rIPC combined with human adipose-derived stem cells (hADSCs) in a rat model. MATERIAL AND METHODS: In total, 24 female Sprague Dawley rats were divided into four groups (n = 6 each): control (skin flap only), rIPC, hADSCs, and rIPC + hADSCs. rIPC was performed in the hind limb of the rats over three cycles of 5 min of occlusion and 5 min of reperfusion, using a tourniquet. A rectangular (3 × 9 cm) dorsal skin flap was used. hADSCs (5 × 105 cells/100 µL) labeled with fluorescent dye were transplanted into the normal subcutaneous tissue at the skin flap boundary. After 14 days, the therapeutic effects of rIPC and hADSCs were evaluated via analysis of the necrotic flap area, histopathologic assessment, and immunohistochemistry (von Willebrand Factor (vWF) and CD31). RESULTS: The necrotic area of the skin flap significantly decreased in the rIPC + hADSCs group (32.75 ± 1.43%) compared with the control (40.60 ± 3.27%, P < 0.01) and rIPC groups (38.84 ± 0.77%, P < 0.05). Dye-labeled hADSCs migrated to the skin flap from the injection site. In the rIPC + hADSCs group, the epithelial tissue and skin appendage had regenerated, and the smooth muscle and subcutaneous fat layers were preserved. Many more vWF- and CD31-positive vessels were observed in the rIPC + hADSCs group compared with the other groups. CONCLUSIONS: The rIPC + hADSCs treatment appeared to reduce skin flap necrosis and activated neovascularization in rats. Therefore, it may be a good strategy for clinical treatment of ischemia-reperfusion injury.
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Precondicionamiento Isquémico , Daño por Reperfusión , Animales , Femenino , Humanos , Necrosis , Ratas , Ratas Sprague-Dawley , Daño por Reperfusión/prevención & control , Células MadreAsunto(s)
Catárticos , Ácido Ascórbico , Colesterol , Pruebas Hematológicas , Humanos , TriglicéridosRESUMEN
BACKGROUND: Automated flow cytometry-based urine analyzer is increasingly being used to identify and enumerate cells and particles in urine specimens. It measures electrical conductivity which could be transformed to osmolality. Using this machine, all urine specimens could be screened for osmolality without requiring a separate dedicated device. We evaluated the performance of the new instrument, the UF-5000 (Sysmex Corporation), in the measurement of urine osmolality. METHODS: The precision of urine osmolality measurement by the UF-5000 was evaluated for 20 days and 4 times a day for 2 concentrations. The linearity and detection capability were evaluated according to the Clinical and Laboratory Standards Institute guidelines. For comparison, 270 random urine specimens from patients were tested simultaneously using the UF5000 and the OsmoPro micro-osmometer (Advanced instruments). RESULTS: The laboratory-based coefficient variations were less than 5%. Urine osmolality using the UF-5000 has a verified linear range (y = 1.097x + 16.91, R2 = .997). Within the comparison analysis, the mean difference was not large (-7.72%) but each differences were largely dispersed with 95% limits of agreement (LoA) from -70.5 to 55.06%, and the mean absolute difference -28.3 mOsm/kg with 95% LoA from -295.13 to 238.45 mOsm/kg. Cohen's kappa value was 0.54 (95% CI, 0.45-0.63). CONCLUSIONS: The UF-5000 measured conductivity and generated an acceptable quantitative analysis of urine osmolality. When compared with the results of the freezing point depression method used by the OsmoPro, a percentage of the measured urine osmolality by the UF-5000 was outside the allowable limit.
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Automatización de Laboratorios , Citometría de Flujo , Urinálisis , Automatización de Laboratorios/métodos , Automatización de Laboratorios/normas , Conductividad Eléctrica , Citometría de Flujo/métodos , Citometría de Flujo/normas , Humanos , Concentración Osmolar , Urinálisis/métodos , Urinálisis/normas , Orina/química , Orina/citologíaRESUMEN
BACKGROUND: We recently introduced the Barricor (BD, Franklin Lakes, NJ, USA) plasma separation tube, which uses a mechanical separator instead of a gel. We evaluated the effects of using the Barricor tube in a stat (statin) laboratory on the results and turnaround time (TAT) of routine chemical tests. We verified the impact of Barricor tube on reducing TAT and providing results similar to those obtained using serum separator tubes (SSTs). METHODS: We collected venous blood samples from 166 outpatients in Barricor tubes and SSTs and measured 28 routine analytes using an AU5800 instrument (Beckman Coulter, Brea, CA, USA). TAT indexes were compared before and after using Barricor tube. RESULTS: Mean percent differences were <5%, except for alanine aminotransferase , total CO2, high-density lipoprotein, phosphate, total protein, and direct bilirubin. The median TAT decreased from 45 to 38 minutes, and the rate of a TAT >60 minutes decreased from 7.84% to 2.66%, which was approximately one-third of that for SST. The reduction in TAT was attributable to a decrease in centrifugation time. Incomplete clotting and repeated centrifugation, which occurred frequently when using SST, also decreased after using the Barricor tubes. CONCLUSIONS: The Barricor tube is an alternative to SST for routine chemical tests in institutions aiming to reduce TAT, with clinically allowable differences in test results.
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Pruebas Diagnósticas de Rutina , Alanina Transaminasa , Instituciones de Atención Ambulatoria , Recolección de Muestras de Sangre , Centrifugación , HumanosRESUMEN
Epigallocatechin-3-gallate (EGCG), the most abundant polyphenol in green tea leaves, has anti-inflammatory effects. In this study, we investigated the mechanism by which EGCG attenuates the effects of lipopolysaccharide (LPS), an agonist of toll-like receptor 4 (TLR4), in cultured human aortic endothelial cells (HAECs). The increase in the expression of intercellular adhesion molecule-1 (ICAM-1) induced by LPS (100â¯ng/ml) was effectively attenuated by pretreatment with EGCG (50⯵M). Importantly, EGCG treatment resulted in a rapid reduction of cellular TLR4, which was accompanied by an increase in the N-terminal fragment of TLR4 in the culture supernatant, indicating that EGCG induces ectodomain shedding of TLR4. EGCG increased cytosolic Ca2+ by inducing the release of intracellular stored Ca2+ and the influx of extracellular Ca2+; accordingly, EGCG-induced ectodomain shedding of TLR4 was nullified by pretreatment with BAPTA-AM (10⯵M), an intracellular Ca2+ chelator. EGCG induced translocation of a disintegrin and metalloprotease 10 (ADAM10) to the cell surface, which was also blocked by BAPTA-AM. Treatment with ADAM10 inhibitor (GI254023X, 2⯵M) and siRNA-mediated depletion of ADAM10 prevented EGCG-induced ectodomain shedding of TLR4 and abolished the inhibitory effect of EGCG on LPS-induced ICAM-1 expression. Collectively, these findings suggest that EGCG decreases cell surface TLR4 in HAECs by inducing ADAM10-mediated ectodomain shedding, and thereby attenuates the effects of LPS. This is a new mechanism of the suppressive effect of EGCG on LPS signaling.